Panicle size is a critical determinant of grain yield in rice and other cereal crops. During rice growth and development, panicle apical abortion (PAA) frequently occurs, particularly under unfavorable conditions, resulting in a reduced number of fertile spikelets and substantial yield losses. In production practice, PAA is a quantitative trait regulated by both genetic and environmental factors. To date, no major quantitative trait loci (QTLs) underlying PAA have been successfully cloned from elite rice cultivars. Studies of PAA-related mutants suggest that this trait involves multiple biological processes. In this review, we synthesize current evidence and propose a pivotal role of calcium in young panicle development. We conclude that PAA is closely associated with disrupted Ca2+ homeostasis and aberrant calcium signaling.
Tillering is a key determinant of rice yield and is regulated by both phytohormone signaling and nitrogen availability. However, how strigolactone-related pathways interface with nitrate-associated processes during tillering remains unclear. Here, we show that the TCP transcription factor OsFC1, a component of strigolactone- and brassinosteroid-associated pathways, exhibits natural promoter haplotypes that are associated with tillering-related variation. OsFC1 overexpression (OsFC1-OE) reduced the transcript abundance of the nitrate transporter gene OsNPF6.3, whereas OsFC1 knockdown (OsFC1-Ri) increased OsNPF6.3 expression. OsFC1 bound to the promoter region of OsNPF6.3 that harbours a TCP-binding motif (GTGGGCCC), and this binding correlated with reduced OsNPF6.3 transcript abundance. Overexpression of OsNPF6.3 in OsFC1-OE plants restored axillary bud elongation and increased tiller number, while knockout of OsNPF6.3 in OsFC1-Ri plants suppressed bud outgrowth and reduced tillering. Together, our findings reveal that OsFC1 acts as a critical molecular link integrating phytohormone signaling and nitrate transport to coordinate rice tillering via affecting OsNPF6.3 transcription.
Rice blast, caused by Magnaporthe oryzae, is the most destructive fungal disease of rice, responsible for annual yield losses of 10–30
Rice sheath blight, caused by the necrotrophic fungus Rhizoctonia solani, poses a major threat to global rice production. Breeding resistant varieties represents the most sustainable management strategy. Cytochrome P450 monooxygenases play diverse roles in plant defense metabolism, including phytoalexin biosynthesis and hormone metabolism. However, their potential involvement in ascorbate-associated resistance pathways remains largely unexplored, and no P450 enzyme has been reported to directly participate in ascorbate biosynthesis in plants. We functionally characterized the rice cytochrome P450 gene OsCYP71D10 through transgenic approaches. Overexpression of OsCYP71D10 in the susceptible variety Lemont significantly enhanced resistance to sheath blight, while RNA interference in the resistant variety Teqing compromised resistance. Integrated transcriptomic and metabolomic profiling of transgenic versus wild-type plants inoculated with R. solani AG1-IA revealed 2252 differentially expressed genes and 25 differentially accumulated metabolites. The “Ascorbate and aldarate metabolism” pathway was the only pathway significantly enriched at both transcriptional and metabolite levels. Notably, L-gulonic-γ-lactone (GulL), an intermediate in ascorbate biosynthesis, showed dramatic accumulation in overexpression lines, accompanied by increased ascorbic acid content and altered expression of redox-related genes (OsGME1, OsAPX, OsDHAR). Exogenous application of GulL and ascorbic acid primed rice immunity against R. solani without direct fungicidal activity, inducing defense-related genes in jasmonic acid and salicylic acid signaling pathways.Our findings establish OsCYP71D10 as a positive regulator of rice sheath blight resistance. Integrated transcriptomic and metabolomic analyses suggest that OsCYP71D10 overexpression is associated with altered ascorbate-related metabolism, redox homeostasis, and defense hormone signaling. However, the direct biochemical function of OsCYP71D10 and its enzymatic substrate remain to be determined. This study provides a novel genetic resource for breeding sheath blight-resistant rice varieties and identifies a previously unrecognized connection between P450-mediated metabolism and ascorbate-related defense responses.
Since most agronomic traits are quantitative, ensuring food security under the pressures of global climate change and population growth increasingly relies on the precise and efficient genetic improvement of crop quantitative traits, making it a central focus of breeding research. Traditional breeding and transgenic approaches are often insufficient for modulating complex quantitative traits. In contrast, the advent of gene-editing technologies has opened new avenues for crop genetic improvement. Notably, the editing of cis-regulatory elements allows fine-tuning of gene expression levels and spatiotemporal patterns without altering coding sequences, enabling targeted optimization of quantitative traits such as yield, quality, and stress resistance. This review systematically summarizes the evolution of cis-regulatory element editing technologies, from early random mutagenesis and screening to targeted dissection guided by functional genomics and, more recently, to intelligent design integrating multi-omics and artificial intelligence, highlighting key technologies, representative applications, and inherent limitations at each stage while discussing future research directions in data integration, algorithm development, and tool deployment. We hope this review will provide both theoretical guidance and practical strategies for intelligent crop breeding.
Seedling cold stress is a major abiotic constraint to rice production, and mining elite cold-tolerant genes from wild rice represents a pivotal strategy to enhance cold tolerance in cultivated rice (Oryza sativa L.). Dongxiang wild rice (DXWR, Oryza rufipogon Griff.) is a valuable genetic resource with robust cold tolerance. However, the underlying molecular regulatory mechanisms remain poorly characterized, and the identification of its elite cold-tolerant genes is still limited. In this study, by integrating high-density gene chip, comparative transcriptomic and functional correlation analyses, we identified OsMYBAS1, an R2R3-MYB transcription factor, as a key regulator conferring cold tolerance of DXWR. The Osmybas1 mutants exhibited drastically reduced survival rate under cold stress, accompanied by excessive reactive oxygen species (ROS) accumulation and significant decreases in antioxidant enzyme activity. Comparative transcriptome analysis of the mutants identified 545 cold-induced differentially expressed genes. Functional enrichment analysis indicated that pathways involved in hormone metabolism and signaling were among the most significantly enriched categories, highlighting their key roles in the cold response. Further detection revealed that endogenous abscisic acid (ABA) and jasmonic acid (JA) levels were markedly down-regulated in Osmybas1 mutants after cold treatment, while exogenous ABA or methyl jasmonate (MeJA) application rescued the cold-sensitive phenotype and reversed the abnormal expression of cold-responsive genes. This study suggested that OsMYBAS1 positively regulated seedling cold tolerance by mediating the coordinated modulation of ABA/JA signaling and ROS homeostasis. These findings elucidated an important molecular mechanism underlying DXWR cold tolerance and provided a novel gene target and theoretical foundation for cold-tolerant rice molecular breeding.
Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5) is encoded by a multigene family in rice (Oryza sativa L.), and its transcriptional abundance is tightly coupled with allelopathic potential. Rice chromosome 2 harbors a tandemly duplicated cluster of four OsPAL paralogs: OsPAL2;1, OsPAL2;2, OsPAL2;3, and OsPAL2;4. To dissect their precise roles in regulating allelopathy, this study generated independent overexpression lines for each OsPAL gene in both the allelopathic rice genotype 'PI312777' and the non-allelopathic cultivar 'Lemont'. Overexpression of individual OsPAL genes significantly enhanced the inhibitory effects of root exudates on barnyardgrass growth, with OsPAL2;1 and OsPAL2;3 exhibiting the most pronounced weed-suppressive phenotypes. Mechanistically, OsPAL overexpression drove distinct tissue-specific metabolic alterations: in transgenic 'PI312777', concentrations of protocatechuic acid, p-coumaric acid, ferulic acid, salicylic acid, and cinnamic acid significantly accumulated in both roots and leaves; conversely, 'Lemont' overexpression lines displayed selective increases in protocatechuic acid, p-hydroxybenzoic acid, and cinnamic acid. Beyond direct allelochemical mediation, OsPAL overexpression reshaped the rhizosphere microbiome. Transgenic 'PI312777' lines displayed reduced alpha diversity and species richness within the root-associated bacterial community. Most strikingly, OsPAL2;1 and OsPAL2;3 overexpression lines showed a marked enrichment of Flavisolibacter, Ohtaekwangia, Lysobacter, and Myxococcota. Collectively, our findings demonstrate that OsPAL2;1 and OsPAL2;3 emerge as prime candidates for engineering next-generation rice varieties with enhanced natural weed-suppressive capacity through integrated metabolic and microbiome engineering.
Rice endosperm, the major edible portion of the grain, plays an important role in regulating blood glucose and preventing intestinal diseases by increasing its resistant starch (RS) content. Previous studies have shown that suppressing amylopectin biosynthesis via genome editing can increase RS content. However, the influence of different Waxy (Wx) allelic backgrounds on RS accumulation in edited lines has not been systematically evaluated. In this study, we used glutinous rice Yunan Heixiangnuo (HXN) with a nonfunctional wx allele and indica rice Yixiang 1B (YX1B) with a weak Wxb allele as backgrounds. We simultaneously knocked out SSSIIIa, SBEI, SBEIIa, and SBEIIb using CRISPR/Cas9, and systematically analyzed changes in RS content, rice quality, and yield traits. The results showed that, in the HXN background, multigene knockout did not significantly alter amylose or RS content but largely maintained favorable eating quality. In contrast, in the YX1B background, quadruple-gene knockout lines exhibited an increase in amylose content from 17.7
Non-structural carbohydrates (NSC) stored in the stem play a crucial role in supporting yield formation in rice. However, internode morphological factors associated with NSC accumulation remain unclear. This study aimed to clarify the relationship between internode morphology and NSC accumulation and to identify a robust morphological indicator for evaluating NSC accumulation capacity. Two years of field experiments were conducted using multiple cultivars. The NSC content was quantified for individual internodes and at the whole-plant culm level, and its relationships with internode morphological traits were analyzed. Since the upper internodes (UIN; first and second internodes) and lower internodes (LIN; third and subsequent internodes) exhibited contrasting roles in NSC accumulation, a novel index was introduced, the volume composition ratio (VCR) of UIN/LIN, which represents their relative volumetric contributions within a culm. The VCR of UIN/LIN showed the strongest correlation with culm NSC and high reproducibility across years, outperforming simple morphological traits. In addition, plant growth regulator treatments that altered VCR were accompanied by changes in culm NSC accumulation. Accordingly, the VCR of UIN/LIN serves as a robust morphological indicator of culm NSC accumulation capacity, providing a practical framework for improving stem carbohydrate storage capacity in rice.
Chalkiness, as a major undesired-index for rice quality evaluation, is the opaque/white structure in rice endosperm, and determines the rice grade and price to a large extent. Reducing chalkiness remains one of the most important goals for breeders and producers. In this study, we identified a rice chalkiness formation-related gene (OsCFF1) that encodes a glycosyltransferase (GT). Through screening of OsCFF1 alleles in 60 rice cultivars, four genotypes were identified, including WT and three natural variations. Cultivars with natural variations that cause the function loss of OsCFF1 usually produce fewer chalky grains. Higher OsCFF1 expression during the grain-filling period is usually associated with a higher degree of chalkiness. Notably, the chalkiness in gene-edited rice kernels was decreased by more than 90
Leaf angle is a key agronomic trait that determines plant architecture and grain yield in rice. Although brassinosteroids (BRs) are known to play a central role in regulating leaf angle, the upstream transcriptional regulatory network remains elusive. Here, we characterized the lla (large leaf angle) mutant which carried a T-DNA insertion in the promoter of OsWRKY11 (a WRKY transcription factor) and exhibits significantly enlarged leaf angles at both seedling and mature stages. We demonstrated that the large leaf angle phenotype of lla was caused by the T-DNA insertion in the promoter of OsWRKY11, leading to its pronounced upregulation. Overexpression of OsWRKY11 recapitulated the lla phenotype, whereas knockdown of OsWRKY11 in the lla background restored leaf angle to wild-type levels, establishing OsWRKY11 as a positive regulator of this trait. Interestingly, the lla mutants are hypersensitive to exogenous BR treatment and accumulate higher levels of the bioactive BR castasterone. Further analyses showed that OsWRKY11 directly binds to W-box elements in the promoter of OsGSR1(a GAST family protein), a known positive regulator of BR biosynthesis, and activates its transcription. Genetic analysis revealed that OsWRKY11 acts upstream of OsGSR1 in the same genetic pathway to regulate leaf angle. Together, our findings uncover an OsWRKY11-OsGSR1 module that fine-tunes leaf angle by modulating BR homeostasis.
Cold stress significantly impairs the rice (Oryza sativa L.) growth and yield, particularly in temperate regions where abrupt temperature fluctuations often occur during the early growth stages. Given the need for novel strategies to improve crop cold tolerance, we evaluated the efficacy of iron oxide nanoparticles (Fe2O3) in enhancing rice cold stress resilience. The reported mechanisms involve promoting plant growth and development, alleviating oxidative stress and inducing defense responses. Using RNA-seq, we analyzed the physiological and transcriptomic responses of rice to cold stress and Fe2O3 treatment. Under cold stress, the NPs elicited a strong antioxidant response-elevating superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities, which led to a marked reduction in oxidative damage, as shown by decreased ROS and MDA levels. Transcriptomic analysis further revealed that NP treatment modulated key pathways related to carbohydrate metabolism, photosynthesis, hormone signaling, and antioxidant metabolism. Collectively, our findings establish that Fe2O3 nanoparticles ameliorate cold stress by preserving chloroplast structure, stomatal architecture, reduce oxidative stress marker, enhancing antioxidant defense system and stabilize photosystem and providing a promising nanozyme-based approach for rice protection against cold induce damage.
Pre-harvest sprouting (PHS), where seeds germinate on panicles before harvest under humid conditions, is a serious global issue in cereal crop production, including rice. Fine-mapping of the previously reported chromosome 4 locus identified OsERF94 as a strong candidate gene for functional validation. In this study, we investigated the role of OsERF94 in PHS using CRISPR/Cas9 gene editing. The CRISPR/Cas9-mediated mutagenesis of OsERF94 induced frameshift mutations, resulting in a loss-of-function of OsERF94 in the 1-I-ET and 2-D-ET lines. The 1-I-ET and 2-D-ET lines exhibited significantly higher germination rates under PHS conditions compared to the wild type, indicating increased susceptibility to PHS. Whole-genome re-sequencing confirmed that few or no mutations could be detected at off-target candidate sites in both edited lines, ensuring the precision of the CRISPR/Cas9 gene editing. A transcriptome analysis revealed altered expression patterns of several GA-related genes, including OsLOL1, OsKO3, OsGA3ox2, and OsGA2ox5 in the OsERF94 mutant lines. The up-regulation of GA biosynthetic genes and the down-regulation of GA deactivation genes observed in both the OsERF94 mutant lines suggest possible alterations in GA metabolism during the early stages of PHS. Transient luciferase reporter assays using a single-luciferase system suggested that OsERF94 may be associated with changes in the promoter activities of several GA- and ethylene-related genes. These findings suggest that OsERF94 may contribute to the regulation of PHS, potentially through moderation of GA- and ethylene-related pathways. Overall, this study improves our understanding of the molecular role of OsERF94 in PHS and highlights its potential as a target for the genetic improvement of PHS resistance in rice-breeding programs.
Saline-alkaline (SA) stress, characterized by high salinity and alkalinity, severely constrains global crop productivity by inducing excessive accumulation of reactive oxygen species (ROS). This study revealed that CRISPR/Cas9-mediated knockout of the NADPH oxidase gene OsRbohD (OsRbohD-KO) significantly enhanced SA stress tolerance in rice. Compared with wild-type plants, the OsRbohD-KO lines exhibited a marked reduction in ROS accumulation in seeds, leaves, and roots under both saline and alkaline stress. This resulted in significantly improved seed germination, bud development, and root elongation. Physiologically, the OsRbohD-KO seedlings maintained higher leaf survival, chlorophyll content and K+ content, while showing reduced malondialdehyde (MDA) levels, membrane injury and Na+ accumulation. In severely SA soils, OsRbohD-KO increased seedling emergence by 5.77–28.85
Grain size and stress resistance are key determinants of high and stable yield in rice. However, the genetic and regulatory mechanisms that coordinate these traits remain elusive. Here, we report that OsHOX24 encoding a HD-ZIP I transcription factor plays a dual role in regulating grain size and salt tolerance. OsHOX24 promotes grain length and weight by enhancing cell division and expansion in spikelet hulls, while it compromises salt tolerance. Additionally, we demonstrated that OsHOX24 participates in ABA signaling and directly activates OsPP2C09 transcription by binding to its promoter, thereby jointly regulate grain size and salt tolerance. Finally, haplotype analysis revealed differentiation of OsHOX24 between indica and japonica, with the prevalent indica haplotype (Hap1) associated with superior grain size and salt tolerance. Taken together, our study not only provides novel insights into the molecular mechanisms that regulate grain size and salt tolerance, but also offers a valuable haplotype-specific target for breeding high-yielding and salt-resilient rice.
The morphology of bulliform phytoliths serves as a key proxy for tracing the origin and evolution of ancient rice cultivation in archaeobotanical research; however, the correlation between these morphological traits and the genetic background of rice is unclear. To the best of our knowledge, for the first time, this study demonstrates that chromosome ploidy level is a key genetic factor influencing rice phytolith morphology, based on a comparative analysis of fan-shaped bulliform phytoliths from haploid and diploid rice. Compared with diploid rice, we found that the bulliform phytoliths in haploid rice exhibited two significant differences: (1) an overall reduction in morphological dimensions (fan width HL = 34.8 μm, fan length VL = 37.6 μm), with dimensions reduced by 18
WRKY transcription factors are important plant regulators that participate in diverse biotic and abiotic stress responses. However, their roles in resistance to Southern rice black-streaked dwarf virus (SRBSDV) remain unknown. Previous work demonstrated that the histone deacetylase gene OsHDA706 (LOC_Os06g37420) enhances SRBSDV resistance by promoting jasmonic acid (JA) biosynthesis. In this study, we first confirmed that the rice line GR216 exhibits strong resistance to SRBSDV through artificial inoculation. Knockout of OsHDA706 in GR216 led to typical SRBSDV symptoms, including dwarfing, dark-green wrinkled leaves, and inverted fibrous roots, confirming its role in antiviral defense. Using the RiceTFtarget database and yeast one-hybrid experiments, we identified OsWRKY30 (LOC_Os08g38990) as a potential upstream regulator of OsHDA706. Co-expression analysis based on the CARFI-Rice database revealed that the expression pattern of OsWRKY30 closely resembles that of several disease-resistance–related WRKY genes under biotic stress and hormone treatments. CRISPR-Cas9 knockout of OsWRKY30 in GR216 resulted in strong symptoms after inoculation with SRBSDV, supporting its involvement in antiviral immunity. RNA sequence (RNA-seq) analysis further showed that OsWRKY30 expression was significantly down-regulated in mutants after SRBSDV infection, and that defense response, stress response, and salicylic acid signaling pathways were significantly enriched in both wild-type and mutant plants. Hormone quantification revealed that JA levels were markedly higher in wild-type than in oswrky30 mutants. Collectively, these results demonstrate that OsWRKY30 positively regulates rice resistance to SRBSDV, likely by modulating JA biosynthesis. This study provides new insights into the molecular mechanisms of rice antiviral defense and offers valuable genetic resources for breeding SRBSDV-resistant cultivars.
The pigmented rice types, i.e., black, red, and purple rice, are highly nutritious substitutes for the ordinary white rice, providing compounds such as anthocyanins, flavonoids, and proanthocyanidins. These bioactive compounds confer antioxidant, anti-inflammatory, anti-cancer, anti-diabetic, and cardiovascular-protective effects, while also assisting in weight management, gut health, and metabolism. The genetic basis of pigmentation includes regulatory genes like OsC1, OsDFR, and OsMYB, which comprise the MYB-bHLH-WD40 complex regulating anthocyanin biosynthesis. Anthocyanin pathways interact with flavonoid and proanthocyanidin synthesis, which is essential for colouration and stress adaptation. The environmental factors, such as temperature, pH, and storage conditions, cause degradation of anthocyanins. Acylation and encapsulation are techniques used in preserving anthocyanins better for industrial use. However, degradation of anthocyanins remains unexplored. Emerging evidence shows that degradation is not only chemical but also enzymatically regulated, mediated by polyphenol oxidases, peroxidases, and β-glucosidases, which accelerate oxidation and hydrolysis reactions and contribute to organ-specific pigment loss. Studies confirm first-order kinetics of anthocyanin breakdown under heat, alkaline pH, and oxygen, while stability improves with encapsulation, co-pigmentation, and gamma irradiation. This review explores the pigmentation of rice (Oryza sativa L.), focusing on its respective health benefits, genetics, synthesis, and degradation. Additionally, much remains to be discovered about its genetic and molecular basis. The regulation of vacuolar enzymes and degradation-related genes in rice tissues is still poorly understood, representing a major knowledge gap compared with biosynthesis.
Seed priming has emerged as an effective and low-cost option to enhance rice seed vigor. However, the fast post-priming deterioration limits its adoption in rice production. The deterioration rate of primed rice seeds was varied across different priming methods, but whether this variation was associated with changes in seed respiratory metabolism remains unknown. In the present study, seeds of rice (Oryza sativa L.) cv. Chuangliangyou627 were subjected to hydro-priming (HP) and osmo-priming with 5
Purple rice has long been treasured for health benefits, but the molecular mechanism of its grain pigmentation remains to be fully understood. Previous evidence shows that a bHLH gene, OsB2, is over-expressed in purple-rice’s caryopsis to cause its increased pigmentation and a MYB gene, OsMYB3, decreases the pigmentation when knocked out in purple rice. Nonetheless, these transcription factors are inadequate to explain why hull color of purple rice can vary independently from that of caryopsis. We show here that more than two regulatory (MYB/bHLH/WDR (MBW)) complexes are involved in grain pigmentation of purple rice. A previously unsuspected MYB, encoded by PURPLE FRUIT 1 (PF1), and OsPa (bHLH), also participate in the grain pigmentation. Along with OsB2 and OsMYB3, they form redundant yet differential MBW complexes in purple-rice grain to regulate not only structural genes of the anthocyanin pathway but also OsLAR of the proanthocyanidin pathway. In the developing caryopses, co-expression patterns of OsPF1 and OsB2 can better predict accumulation process of anthocyanins than those of OsMYB3 and OsB2. Disrupting OsPF1 or OsPa via CRISPR/Cas9 in purple rice leads to reduced pigmentations of caryopsis and hulls, with OsPa’s knockout-effect on hull color particularly pronouncing. When paired with OsPa and OsTTG1 to form the MBW complex, OsPF1 can initiate a stronger activation of OsCHI but weaker one of OsLAR than OsMYB3 under the same condition. Compared to significantly enhanced expression levels of OsB2 and OsPa in grains of purple rice, transcriptions of OsPF1 and OsMYB3 remain non-differentiated between straw-white and purple grains at the same developmental stage. The new advances, along with the previously known, have enabled a mechanistic illustration of pigmentations of caryopsis and hulls in purple rice, which may guide future breeding of purple rice in creating varieties of desired grain anthocyanins to improve human health.
